Quantitative analysis of cells and analytes in fluid samples, particularly bodily fluid samples, often provides critical diagnostic and treatment information for physicians and patients. Immunological testing methods (Kennedy, D. M. and S. J. Challacombe, eds., ELISA and Other Solid Phase Immunoassays: Theoretical and Practical Aspects, John Wiley and Sons, Chichester (1988)), which take advantage of the high specificity of the antigen-antibody reaction, provide one approach to measurement of analytes. Immunoassays which provide a quantitative measurement of the amount of an analyte in a sample often use complex, multistep procedures and expensive analyzers available only in a laboratory setting. Immunochromatographic assays, such as those described in GB 2,204,398A; U.S. Pat. Nos. 5,096,837, 5,238,652, and 5,266,497; Birnbaum, S. et al., Analytical Biochem. 206:168-171 (1992); Roberts, M. A. and R. A. Durst, Analytical Chem. 67:482-491 (1995); and Klimov, A. D. et al., Clinical Chem. 41:1360 (1995), are simpler. Immunochromatographic assays rely for their interpretation on the observation of colored reaction products, usually colored particles, in a particular region of a membrane. To obtain a quantitative measure of the amount of an analyte detected in an immunochromatographic assay, the amount of colored particles is analyzed.
Measurement by optical means of the amount of particles accumulated in one or more regions of a membrane is usually performed by reflectometry or densitometry. In these techniques, a light beam is either reflected or transmitted through the colored portion of the membrane, and the light intensity reaching a detector is measured. Other methods include obtaining, with a video camera, the image of the membrane illuminated with transmitted or reflected light, and performing image analysis on a frame of the video record. Although image analysis software allows considerable manipulation of the image to optimize sensitivity in detection of the particles, these methods suffer from light scattering by the membrane fibers in which the particles are imbedded. The multiple scattering that results obscures the optical signal produced by the particles and dramatically increases the noise level in the measurement.